ES2823001T3 - Sealing of reaction cells for bioaffinity assays - Google Patents

Abstract

System (20) comprising a bioassay cartridge (4), comprising reaction chambers (6) containing bioaffinity reagents in a dry state, and a pierceable hermetic cover (2) that does not allow, before being pierced, any flow or diffusion of matter to or from said reaction chamber (6) through said cover (2), characterized in that: a) said cover (2) comprises at least a first layer (8), that is, a layer upper layer (8), a second layer (10), that is, an intermediate layer (10), a third layer (12), that is, a lower layer (12), and sites intended for perforation (14), b ) in the event that said cartridge (4) is covered with said cover (2), said third layer (12) is against said cartridge (4) and said sites (14) intended for perforation are in openings (16 ) of the reaction chambers (6), c) said cover (2) has, at the sites (14) intended for drilling, a hollow space (18) between said first ca pa (8) and said third layer (12), that is, said second layer (10) has a hole (18) that extends through said second layer (10), and d) the first layer (8) or the third layer (12), preferably said first layer (8), of the cover (2) is hermetic until perforation, and the third layer (12) or the first layer (8), respectively, preferably said third layer (12) , is pre-grooved so that: i) in the event that it is pierced with a needle, the piercing point is not gas-tight, but allows the free flow of gases out of the reaction chamber (6) , and ii) said layer guarantees a hermetic closure of the needle channel with the retraction of said needle, in which the cover (2) comprises an additional layer (22) in the upper part, that is, on, the first layer (8), and said additional layer (22) has, at the places (14) intended for drilling, a hollow space (24).

Description

DESCRIPTION

Sealing of reaction cells for bioaffinity assays

Field of the invention

The invention relates to in vitro diagnostic tests of analytes from biological or clinical samples. In greater detail, the invention relates to patient-friendly in vitro diagnostic assays of clinical samples applying bioaffinity binding reactions. In particular, the invention relates to the sealing of reaction cells containing dry reagents for bioaffinity assays.

Background of the invention

Publications and other materials used herein to illustrate the background of the invention, and in particular cases to provide additional information regarding practice, are incorporated by reference. Trends in diagnostic testing

A wide variety of methods and instruments are commercially available for in vitro immunodiagnostic (IVD) assays of clinical specimens. Traditional IVD assays, such as ELISA immunoassay assays, are characterized by complex assay methodology. An assay may require the addition of reagents in several stages and washing in several stages. This causes the tests to be laborious in execution. In order to reduce the effort required, automated analyzers have been developed. The analyzers can operate in "random access mode" or in "batch mode". Automated analyzers can perform up to several hundred tests per hour. Typically, the larger the analyzer, the greater the assay capacity. The assay menu of an automated random access analyzer can contain assays for up to 50 different analytes, or even more. For economy of scale, a large analyzer can provide more economical results than a small analyzer. This has driven IVD testing to large centralized laboratories.

The main disadvantage of centralized trials is the long turnaround time, which is excessively long to meet the testing needs of acute patients. Therefore, the trend of centralization has been followed by the trend of trials close to the patient, that is, the trial at the site of care. At the site of care, there is a growing need for test instruments that provide rapid results. To be applicable at the site of care, the instrument should be easy to use, small in size and affordable in cost.

In order to meet the requirements for on-site testing, the testing methodology should be as simple as possible. A widely used approach to simplifying assay methodology is to apply dry (or lyophilized) biochemical reagents instead of liquid reagents. Using dry reagents can eliminate reagent addition steps.

Another approach to simplify the assay methodology is to apply a detection technology that allows non-separation (washout) detection of bioaffinity assays. Using a separation-free detection technique can eliminate washing steps.

One approach to reducing the size of the analyzer is to reduce reaction volumes, that is, to miniaturize the assay system. This also reduces the volumes of assay consumables, such as assay reagents and buffers. This makes the test more suitable for use at the point of care. However, miniaturization usually compromises the performance figures of the sensing technique. To avoid the above, a detection technique should be used that tolerates miniaturization without compromising performance.

Dry reagents

It is widely known that bioaffinity reagents, such as antibodies, antigens and enzymes, retain biological activity very well in the dry state. In the dry condition, the reagents are usually stable on storage even at room temperature. In this way, there is no need to maintain a strict cold chain in reagent supply logistics. This reduces transportation and storage costs. The dry reagents also allow the design of simpler test instruments for use at the point of care.

It is also common knowledge that dry bioaffinity reagents must be kept tightly closed to avoid contact with ambient moisture. Upon exposure to moisture, dry reagents tend to lose biological activity, leading to reduced assay performance. In the event that the assay reagents dry out in the final reaction cell, the reaction cell must be hermetically sealed to prevent contamination. contact with humidity. Most often, this is done with an adhesive metal sheet. To improve the mechanical properties, the sheet can be composed of several coats of varying materials. A common type of foil is made up of a plastic layer and a metal foil layer. The plastic layer allows the blade to be more durable and flexible. In the event that hermetic sealing is not required, the reaction cell can be sealed with a bare plastic film to protect it from dust and other occasional spills.

In a typical automated IVD analyzer using dry reagents, the clinical sample can be dispensed through the cover sheet into the reaction cell using a dispensing needle. The dispensed sample dissolves the dry reagents and induces the binding reaction between the analyte and the reagents. Mixing or agitation of the reaction cell is often necessary to accelerate the dissolution of the reagents and enhance the reaction kinetics. In point-of-care contexts, fast reaction kinetics are essential due to the need for a short response time. In most analyzers, post-processing of the reaction well is typically required, such as washing unbound components and adding components that allow quantification of the degree of immunoassay binding (e.g., substrate or solution intensifier). Thus, access to the well is required several times.

Shaking open reaction cells tends to cause spillage and aerosol formation, which can lead to contamination of nearby reaction cells. This can cause false test results and deteriorate both the accuracy and precision of the test method. In this way, mechanical mixing is associated with a significant risk of cross contamination.

In the case of miniaturized test systems where the reaction volume is small, evaporation of the solvent from an open cuvette could also play a role to a significant degree. In such a case, the actual concentrations increase, distorting the test results. In miniaturized systems, the effects of spillage and aerosol formation are pronounced compared to conventional sized buckets.

Evaporation and spillage caused by agitation could be avoided by sealing the test cells after dispensing the sample. However, sealing the cuvettes would complicate the manual assay protocol or, if the method is automated, significantly complicate the analyzer design. In conclusion, a sealing step should be avoided to make the analyzer suitable for routine IVD used at the point of care.

In the event that the cuvette is covered with a sheet (or other type of cover) and the dispensing of the samples is carried out through the sheet with a thin dispensing needle, the probability of spillage would be reduced compared to the open cuvettes. In this case, the probability of leakage would be proportional to the diameter of the piercing needle. However, even in this case, spillage will most likely occur during shaking and significant evaporation is likely to occur during incubation. This may deteriorate the performance of the assay.

Resealable pierce covers

In order to overcome the problems listed above, the trays could be sealed with a resealable pierceable cover. Many types of resealable covers are known in the art. Said covers can be made of plastic films or flexible materials, such as rubber, silicone and other elastomers. Such covers are widely applied to cover, for example, reaction vials of nucleic acid amplification reactions, such as thermocycled PCR reactions. In these, the seal is typically pierced after cycling to aspirate the liquid. However, such covers are difficult to apply in miniaturized reaction cells, such as the microtiter wells of the 384-well format. One of the main obstacles to such elastomer covers is the increase in air pressure in the bucket due to dispensing. In order to avoid increased pressure, an equivalent volume of air should flow out of the cuvette. In the case of a rubber or silicone cup, the dispensing needle sits firmly in the perforated opening and does not let air flow out. Increased pressure impairs dispensing accuracy or may prevent dispensing altogether. In conclusion, pierceable covers made of molded rubber, silicone or other resilient / elastic bulk material are not suitable for covering small volume reaction cells.

Increased pressure problems can be overcome by pre-gouging (pre-cracking) the sealing material at the expected point of piercing. Pre-grooving can be linear, Y-shaped, cross-shaped, or otherwise. After piercing with a needle, the edges of the slot would bend downward, thus opening a cutout for the free exit to the outside of air. Upon retraction of the needle, the edges must revert to their original position for proper closure of the opening. Therefore, the cover material must be elastic and / or resilient. The complete pre-grooving of the cover material allows the free diffusion of ambient gases into the cuvette, thus the closure is not hermetic. Accordingly, fully pregrooved sealants are not applicable without modification to dry reagents. The elastic cover, whether pre-slotted or not, can be covered with a metallic layer to keep the cover watertight until pierced with a needle. Such cover materials are commonly used to bag microtiter plates, strips, and other bioassay-sensitive bioassay consumables. humidity. However, the metallic layer is inelastic. In this way, it resists the bending of the edges of the slit. Once the edges are bent down through the perforation, the metal layer offers resistance to the edges returning to their original position. In other words, the metal foil disturbs the correct reversible function of the pre-grooved elastomer cover. The case of an incorrect closing of the opening can lead to spillage or evaporation of the reaction mixture. This again deteriorates the performance of the method.

US 2008/152545 discloses a kit containing a specimen retrieval device. Document No. US 2004/0235036 discloses devices and methods for detecting genetic sequences. Document Nos. FR2938063A1 and US5789251 disclose cartridges with a plurality of chambers. US2006 / 226113A1 and US2008 / 251490A1 disclose specimens with sealing caps.

None of the prior art methods for sealing reaction cells satisfy the criteria of: (i) tightness during storage,

(ii) allow accurate dispensing with a pierced needle,

(iii) allow air outflow during dispensing,

(iv) reversibly close the perforated opening to prevent spillage and evaporation.

Objective and summary description of the invention

An object of the present invention is to provide a system comprising a bioassay cartridge with reaction chambers and a cover for said cartridge.

The present invention provides a system comprising a bioassay cartridge comprising reaction chambers and a pierceable hermetic cover for said cartridge. The characteristics of the cover are that: a) said cover comprises at least a first layer, that is, a top layer, a second layer, that is, an intermediate layer, a third layer, that is, a bottom layer, and sites intended for perforation, b) in the event that said cartridge is covered with said cover, said third layer is against said cartridge and said sites intended for perforation are in openings of the reaction chambers, c) said The cover has, at the places intended for the perforation, a hollow space between said first layer and said third layer, that is, said second layer has a hole that extends through said second layer, and

d) the first layer or the third layer, preferably said first layer, of the cover is hermetic until perforation, and the third layer or the first layer, respectively, preferably said third layer, is pre-grooved so that:

i) In the event that it is pierced with a needle, the piercing connector is not gas-tight, but allows free flow out of gases from the reaction chamber, and

ii) said layer guarantees a hermetic closure of the needle channel with the retraction of said needle, in which the cover comprises an additional layer in the upper part, that is to say, over, the first layer, and said additional layer presents, in the sites intended for drilling, a hollow space.

Brief description of the drawings

Figure 1 shows schematically, with an exploded view of the cover, a single-well bioassay cartridge system not according to the invention.

Figure 2 shows schematically, with an exploded view of the cover, a 12-well bioassay cartridge system not according to the invention.

Figure 3 shows schematically, with an exploded view of the cover, a 96-well bioassay cartridge system not according to the invention.

Figure 4 shows schematically, with an exploded view of the cover, a 384-well bioassay cartridge system not according to the invention.

Figure 5 shows schematically, with an exploded view of the cover, another 384-well bioassay cartridge system not according to the invention.

Figure 6 shows schematically, with an exploded view of the cover, a further system of 384-well bioassay cartridges according to the invention.

Figure 7 shows schematically, with an exploded view of the cover, a 384-well bioassay cartridge system according to the prior art.

Detailed description of the invention

The invention provides a new design for sealing low volume bioaffinity test cartridges. The design is particularly suitable for random access analyzer assays where samples that must be dispensed into one or more parallel reaction chambers are inserted at irregular intervals for analysis and it is important that reaction chambers to be used later are kept tight. The new design enables the manufacture of ready-to-use bioassay cartridges with reduced volume reaction chambers, which:

(i) contain bioaffinity reagents in a dry state,

(ii) are kept hermetically closed during storage,

(iii) allow accurate dispensing into the chamber with a piercing needle,

(iv) allow a free outflow of air from the chamber during dispensing,

(v) guarantee reversible closure of the needle channel with retraction,

(vi) eliminate cross contamination caused by occasional spills.

Typical features of the new seal design are as follows:

(i) the seal has a pre-grooved bottom layer made of resilient material,

(ii) the seal has a tight top layer, and

(iii) the seal has a hollow / spacious interlayer.

The hollow interlayer is the essence of the invention. A seal in accordance with the present invention overcomes the obstacles of the prior art and allows the manufacture of ready-to-use low volume bioassay cartridges that satisfy the four criteria listed above.

According to the invention, a hollow intermediate layer separates the lower layer from the upper layer. The middle layer provides space between the top and bottom layers and keeps the two layers at an essentially constant distance from each other.

The hollow interlayer is essential for the proper function of the cover. Without the hollow interlayer, the cover does not meet the imperative requirements for ready-to-use low-volume bioassay cartridges. The structure of a typical roof not according to the invention is shown in figure 1. Figure 1 shows a side projection. Figure 1b shows a plan projection. The thickness of the hollow layer is typically at least 0.2mm. The preferred thickness is at least 0.5mm. In the event that the thickness is excessively reduced, the layer gradually loses its effect of resistance to the consequences of spillage. In principle there is no maximum thickness for the intermediate layer. However, for practical reasons, a preferred thickness is at most 10mm. The most preferred thickness is 1 to 5mm.

The middle layer is hollow at the point of perforation. The hollow space may be in the shape of a truncated cylinder, cone, cone or cube, or any other shape. The volume of the hollow space is proportional to the thickness of the layer and depends on the shape of the hollow space. Typically, the volume is not less than 5% of the volume of the cartridge cavity, that is, the reaction chamber. In the event that the volume is excessively small, the layer loses its effect of resistance to the consequences of spillage and the ability to allow the free operation of the bottom and top layers. There is no upper limit to the volume of the space, although for practical reasons the volume should not exceed the volume of the cartridge cavity by more than 10 times.

The hollow middle layer is attached on the upper face of the upper layer. The top layer can be any material that is pierceable with a needle and is watertight until pierced. After drilling it is no longer airtight. The upper layer can be made of metal foil or plastic-metal bilayer or other composition. The composition and dimensions of the top layer do not limit the scope of the invention.

The hollow intermediate layer is attached inferiorly to the lower layer. The bottom layer is made of any elastic or flexible material that is pierceable with a needle and allows the outflow of air from the cartridge during dispensing. The bottom layer is pre-grooved before drilling. The lower layer can be composed of any elastic or flexible material, such as plastic film, cellular foam, polyurethane, rubber, silicone or other material, provided that, when pierced with a needle, the piercing connector is not hermetic. to air, but allows free flow of air out from the cartridge cavity.

Terms of use

The terms used in the present application can be defined as follows:

• Pierceable seal: In the context of the present invention, the term “ pierceable seal” refers to a bioassay cartridge cover which is called cartridge reaction chambers. The reference to the cover being hermetic means that the cover, prior to being pierced, does not allow any flow or diffusion of matter to or from a reaction chamber through the cover. Accordingly, in the context of the present application, the hermetic cover ensures that the dry reagents, typically dried or lyophilized, do not deteriorate due to the flow or diffusion of matter, typically water vapor, into the chamber. from reaction through the shell, not even during prolonged storage, that is, storage lasting at least several weeks, preferably months. Reference to pierceable means that the cover can be pierced with a dispensing needle for sample insertion and optionally a dilution buffer in conjunction with, and / or in addition to, reagents.

Bioassay Cartridges: In the context of the present invention, the term " bioassay cartridge" refers to any cartridge, be it a single tube, a multi-reaction well strip (e.g. 12 wells) or a multiwell plate (e.g. ., 96 or 384 wells). In the context of the present application, the term typically refers to cartridges for bioassays in which the volume of the reaction chambers is 5 µl to 2 ml, preferably 5 µl to 50 µl, 50 µl to 500 µl, or 500 µl to 2 ml, and most preferably 10 µl to 30 µl.

First layer / Top layer: in the context of the present invention, the reference to first layer and top layer of the cover of the bioassay cartridge refers to the layer of the cover that is on top of the other layers defined in the application, it is that is, the layer that is on the middle layer, which is on the lower layer, of the cover at the time the cover seals the cartridge.

Second layer / middle layer : In the context of the present invention, the reference to second layer and middle layer of the bioassay cartridge cover refers to the layer of the cover that lies between the upper layer and the lower layer of the cover. It should be noted that the intermediate layer of the cover can be a continuation of the upper and / or lower layer, provided that an intermediate layer can be defined, between the upper layer and the lower layer, so that the intermediate layer comprises a hollow space or hollow spaces between said first layer and said third layer, that is, said second layer has a hole or holes that extend through said second layer at the site or sites, respectively, intended for perforation. Third layer / Lower layer, in the context of the present invention, the reference to third layer and lower layer of the cover of the bioassay cartridge refers to the layer of those defined in the invention, which is against, that is, more close to the reaction chamber, in particular the opening of the reaction chamber at the time the cartridge is covered with the cover, eg sealed with the cover.

Site / Sites intended for drilling: in the context of the present invention, the reference to site intended for drilling and sites intended for drilling refers to sites, that is, particular areas, of the deck surface or surface of a particular layer of the cover of the bioassay cartridge through which the perforation is carried out for the insertion of sample and optionally of a buffer for dilution together with, and / or in addition to, reagents, when the cartridge is used, that is, when the bioassay is carried out. The site or sites for perforation are at the opening of the reaction chamber or the openings of the reaction chambers of the bioassay cartridge at the time the cartridge is covered with said cover, e.g. at the time it is sealed with the cover.

Hollow space / thickness of the hollow space / width of the hollow space: in the context of the present invention, the term hollow space refers to the holes in the intermediate layer of the cover of bioassay cartridges. The hole extends through the second layer from the first layer to the third layer. Accordingly, the holes are limited by the upper layer, superiorly, the intermediate layer on the sides, and the lower layer, inferiorly. The term " gap thickness" refers to the distance between the first layer and the second layer along the gap. Thickness is typically measured parallel to the desired drilling axis. The desired drilling axis is typically perpendicular to the plane of the deck. The thickness of the hollow space is equal to the thickness of the intermediate layer in the case that the thickness of the intermediate layer is constant, which is preferably the case. The term " gap width" refers to the dimension of the gap perpendicular to the desired drilling axis and typically parallel to the plane of the deck. The width of the hollow space can vary in relation to the distance between the upper layer and / or the lower layer depending on the shape of the hollow space. In the case where the shape is eg that of a cone or truncated cone, the width of the hollow space will depend on at which end the cone or truncated cone is measured.

Reaction chamber / reaction chamber volume: in the context of the present invention, the term reaction chamber refers to the space limited by the walls of the reaction chamber, typically the tube or well, and the plane of the cover that covers the bioassay cartridge. Accordingly, the volume of the reaction chamber refers to the total volume of the chamber in which the bioassay reaction must take place. In this way, the volume is also limited by the walls of the reaction chamber, typically the tube or well, and the plane of the cover that covers the bioassay cartridge. Typical volumes of the reaction chamber of the present invention are between 5 µl and 500 µl, preferably between 5 µl and 50 µl or between 50 µl and 500 µl, and most preferably between 10 µl and 30 µl.

Piercing connector: In the context of the present invention, the term piercing connector refers to the needle connector that has pierced the sheath or a particular layer of the sheath. Typically, the piercing connector through the upper layer or the lower layer or both, preferably at least the lower layer, is not gas-tight, but allows free flow out of gases from the reaction chamber. by dispensing the sample and optionally a dilution buffer together with, and / or in addition to, reagents, into the reaction chamber.

Needle channel / needle channel seal: In the context of the present invention, the term " needle channel" refers to the channel through the cover or a particular layer of the cover left by the piercing needle after it is has retracted. Typically, at least the needle channel through the upper layer or the lower layer is sealed upon retraction of the needle. The term " hermetically sealed" in the context of the present invention means that the closure does not allow a seal to occur. significant flow of matter, that is, flow of matter, that can significantly affect the performance of the bioassay performed, through the needle channel that is hermetically sealed during the bioassay. Preferred embodiments of the invention

A system of the definition is defined in claim 1. It comprises, inter alia, a pierceable hermetic cover for a bioassay cartridge with reaction chambers, wherein:

a) said covering comprises at least a first layer, that is, a top layer, a second layer, that is, an intermediate layer, a third layer, that is, a bottom layer, and sites intended for perforation, b ) in the event that said cartridge is covered with said cover, said third layer is against said cartridge and said sites intended for perforation are in openings of the reaction chambers, and c) said cover has, in the places intended to the perforation, a hollow space between said first layer and said third layer, that is, said second layer has a hole extending through said second layer.

The cover, before being perforated, does not allow any flow or diffusion of matter to or from a reaction chamber through the cover.

In most typical embodiments of the present invention, the volume of each void at each perforation site is between 5% and 10 times, preferably between 15% and 3 times, and most preferably between 50% and 2 times. the corresponding reaction chamber volume of the cartridge. In many typical embodiments, the thickness of the void, that is, the distance between the first layer and the second layer through the void, is between 0.1 mm and 20 mm, preferably between 0.3 mm and 10 mm. mm, and most preferably between 1 mm and 5 mm, and / or the width, measured essentially perpendicular to the desired drilling axis, of the hollow space at the drilling site is between 1.5 mm and 2 times, preferably between 2 mm and 1.5 times, and most preferably between 2.5 mm and 1 times the width of the opening of the reaction chamber covered with said cover.

In a system of the invention, the first layer or the third layer, preferably said first layer, of the cover is hermetic until perforation, and the third layer or the first layer, respectively, preferably said third layer, is such that:

i) in the event that it is pierced with a needle, the piercing point is not gas-tight, but allows the free flow out of gases from the reaction chamber, and

ii) said layer guarantees a hermetic closure of the needle channel with the retraction of said needle.

The layer, be it the first layer or the third layer, preferably said first layer, with a piercing connector that is not gas-tight, when pierced by a needle, although it allows the free flow of gases out of the chamber, it is pre-grooved. Preferably, the pre-groove is in the shape of (ie, cross-shaped), X-shaped, Y-shaped, or I-shaped (ie, linear).

In some preferred embodiments of the invention, the cover comprises at least one additional layer. The additional layer or layers may be on, between or under the first, second and / or third layers. In a system of the invention, the cover comprises an additional layer in the upper part, that is to say, over, the first layer, and said additional layer has, at the places intended for the perforation, a hollow space.

A typical system according to the invention comprises a bioassay cartridge with reaction chambers and a cover for said cartridge, wherein the cover is as defined above. In most typical embodiments of the system, the volumes in the reaction chambers of the bioassay cartridge are between 5 and 2 ml, preferably between 5 and 50 ml, between 50 and 500 ml, or between 500 ml. and 2 ml, and most preferably between 10 µl and 30 µl.

In most typical use embodiments, the volumes of the reaction chambers of the bioassay cartridge are between 5 and 2 ml, preferably between 5 and 50 ml, between 50 and 500 ml, or between 500 pl and 2 ml, and most preferably between 10 and 30 ml.

Examples

The invention is illustrated by Examples 1 through 7, below; however, the applications in which the present invention provides advantages are not limited to the present examples.

Example 1 (not according to the invention)

Single Well Reaction Chamber

Figure 1 shows a bioassay cartridge 4 with a single-well reaction chamber 6 sealed with a three-layer cover 2 8, 10, 12. The lower layer 12 of cover 2 is made of 3 mm thick silicon , pre-grooved (X shape) at the intended drilling point. The hollow space 18 of the intermediate layer 10 is cylindrical in shape, 10 mm in diameter and 10 mm deep. The lower layer 10, that is, the skeleton around the hollow space 18, which joins the upper layer 8 and the lower layer 12, is made of closed-cell polyethylene foam. The upper layer 8 is hermetic, made of metal sheet, 80 µm thick. Tube 4 is filled with dry reagents. The reagent cartridge 4 is stored in a foil pouch until it is used for the assay. Cartridge 4 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The three-layer cover 2 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the essential advantages of the invention.

Example 2 (not according to the invention)

Multiwell cartridge, 12 reaction wells

Figure 2 shows a system 20 comprising a multiwell cartridge 4 composed of 12 reaction wells 6 in a sealed matrix with a three-layer cover 2 8, 10, 12. The lower layer 12 of cover 2 is made of a foam 2mm closed cell neoprene, pre-grooved (Y-shape) at the intended perforation point. The hollow space 18 of the intermediate layer 10 is cuboid in shape (6mm x 6mm) and 2mm deep. The intermediate layer 10, that is, the skeleton around the hollow space 18, which joins the upper layer 8 and the lower layer 12, is made of closed-cell foam rubber. The upper layer 8 is hermetic, made of metal laminated with plastic (bilayer) with a thickness of 120 µm. Reaction chambers 6 are filled with dry reagents.

Cartridge 4 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The three-layer cover 2 8, 10, 12 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the essential advantages of the invention .

Example 3 (not according to the invention)

Multiwell cartridge, 96 reaction wells

Figure 3 shows a system 20 comprising a multi-well cartridge 4 composed of 96 reaction wells 6 constituted by a standard 96-well plate 20, in which the cartridge 4 is sealed with a three-layer cover 2 8, 10, 12 The lower layer 12 of the cover 2 is made of 100 µm thick vinyl, pre-grooved (form I) at the intended perforation point. The hollow space 18 of the intermediate layer 10 is conical in shape, 5 mm in diameter and 1 mm deep. The intermediate layer 10, that is to say the skeleton around the hollow space 18, is made of polyurethane. The upper layer 8 is hermetic, made of metal sheet, 15 µm thick. Cartridge system 20 is filled with dry reagents. The reagent cartridge system 20 is stored in a foil pouch until used for the assay.

The cartridge system 20 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The three-layer cover 2 8, 10, 12 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the essential advantages of the invention .

Example 4 (not according to the invention)

Multiwell cartridge, 384 individual reaction wells

Figure 4 shows a multiwell cartridge system 20 composed of 384 individual reaction chambers 6 made in a standard 384-well plate 4 sealed with a three-layer cover 2 8, 10, 12. The bottom layer 12 of the cover is hermetic , made of metal sheet 50 pm thick; the upper layer 8, made of polyurethane cell foam, is pre-grooved (shaped) at the intended perforation point 14 and is 0.5 mm thick. The metallic layer 12 is not pre-grooved. The hollow space 18 of the intermediate layer 10 is cylindrical in shape, 2 mm in diameter and 0.5 mm deep. The intermediate layer 10, that is, the skeleton around the hollow space 18, is made of closed-cell foam. System 20 is filled with dry reagents. The cartridge system 20 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The three-layer cover 2 8, 10, 12 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the advantages of the invention.

Example 5 (not according to the invention)

Multiwell cartridge, 384 individual reaction wells

Figure 5 shows a multiwell cartridge system 20 composed of 384 individual reaction chambers 6 made in a standard 384-well plate 4 sealed with a three-layer cover 2 8, 10, 12. The bottom layer 12 of cover 2 is made of a 300 µm polyethylene-closed cell polyurethane foam bilayer, pre-grooved (shaped) at the intended perforation point. The hollow space 18 of the intermediate layer 10 is cylindrical in shape, 3 mm in diameter and 2 mm deep. The intermediate layer 10, that is, the skeleton around the hollow space 18, is made of closed-cell foam. The upper layer 8 is hermetic, made of aluminum sheet, 30 µm thick. System 20 is filled with dry reagents.

The cartridge system 20 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The three-layer cover 2 8, 10, 12 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the essential advantages of the invention .

Example (according to the invention)

Multiwell cartridge, 384 individual reaction wells

Figure 6 shows a multiwell cartridge system 20 otherwise identical to Example 5, but features an additional layer 22 similar to intermediate layer 10 on top layer 8. Additional layer 22 may, in some embodiments, improve performance. through more efficient segregation of the sites intended to be drilled. In this way, in the event of a spill at the drilling site, the risk of the spill contaminating other drilling sites is greatly reduced.

The cartridge system 20 is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. The four-layer cover 2 22, 8, 10, 12 is punctured with the needle, dispensing the sample volume into reaction chamber 6 and then retracted from chamber 6. The design of said cover 2 provides the essential advantages of the invention.

Example 7 (not according to the invention)

Multiwell cartridge, 384 individual reaction wells

Figure 7 shows a prior art multiwell cartridge system 20 'composed of 384 individual reaction chambers 6 made in a standard 384-well plate 4 sealed with a standard cover material 2' made in a foil bilayer 8 - plastic 12. The plastic layer 12 (at the bottom) is pre-grooved (shaped) at the intended piercing point. The upper layer 8 is hermetic, made of metal foil. Reaction chambers 6 are filled with dry reagents.

The cartridge system 20 'is used for a bioassay. A sample is added to reaction chamber 6 with a dispensing needle. When piercing the bilayer cover 2 'with a needle, the edges of the pre-grooved layer 12 are bent inwards, while in the retraction of the needle, the edges do not reverse correctly because the metal foil layer 8 is not sufficiently elastic. . Thus, a sufficient seal of well 6 is not achieved after sample addition. Furthermore, the close proximity of the pre-grooved layers 12 and hermetic 8 wrap around the dispensing needle with excessive force to allow the replacement air to flow out reliably. Furthermore, the design is vulnerable to contamination from well 6 to well 6 'due to spillage. The design of said cover 2 'represents the state of the art. The hollow layer is missing; thus, such a cover does not provide the advantages of the invention.

In the event that the pre-grooved plastic layer was at the top and the foil was at the bottom, an additional problem would be the occasional chunks of foil falling into the reaction chambers at the reaction sites. drilling.

Claims (7)

1. System (20) comprising a bioassay cartridge (4), comprising reaction chambers (6) containing bioaffinity reagents in a dry state, and a pierceable hermetic cover (2) that does not allow, before being pierced , any flow or diffusion of matter to or from said reaction chamber (6) through said cover (2), characterized in that: a) said cover (2) comprises at least a first layer (8), that is, an upper layer (8), a second layer (10), that is, an intermediate layer (10), a third layer (12 ), that is, a lower layer (12), and sites intended for drilling (14), b) in the event that said cartridge (4) is covered with said cover (2), said third layer (12) is against said cartridge (4) and said sites (14) intended for perforation are in openings ( 16) of the reaction chambers (6), c) said cover (2) has, at the sites (14) intended for drilling, a hollow space (18) between said first layer (8) and said third layer (12), that is, said second layer (10) has a hole (18) that extends through said second layer (10), and d) the first layer (8) or the third layer (12), preferably said first layer (8), of the cover (2) is hermetic until the perforation, and the third layer (12) or the first layer (8) , respectively, preferably said third layer (12), is pre-grooved so that: i) in the event that it is pierced with a needle, the piercing point is not gas-tight, but allows the free flow of gases out from the reaction chamber (6), and ii) said layer guarantees a sealing of the needle channel with the retraction of said needle, in which the cover (2) comprises an additional layer (22) in the upper part, that is to say, over, the first layer (8), and said additional layer (22) has, at the sites (14) intended for drilling, a hollow space (24). System (20) according to claim 1, characterized in that, at each perforation site (14), the volume of each hollow space (18) at each perforation site is between 5% and 10 times, preferably between 15% and 3 times, and most preferably between 50% and 2 times the volume of the corresponding reaction chamber (6) of the cartridge (4). System (20) according to claim 1 or 2, characterized in that the thickness of the hollow space (18), that is, the distance between the first layer (8) and the second layer (12) through the hollow space (18 ), is between 0.1 mm and 20 mm, preferably between 0.3 mm and 10 mm, and most preferably between 1 mm and 5 mm. System (20) according to any one of claims 1 to 3, characterized in that the width, measured essentially perpendicular to the intended drilling axis, of the hollow space (18) at the drilling site (14) is between 1.5 mm and 2 times, preferably between 2 mm and 1.5 times and most preferably between 2.5 mm and 1 times the width of the opening of the reaction chamber (6) covered with said cover (2). System (20) according to any of the preceding claims, characterized in that the layer, be it the first layer or the third layer, preferably said first layer, with a piercing connector that is not gas-tight, as it is pierced by a Although the needle allows free flow of gases from the chamber, it is pre-grooved in the shape of an X, Y-shaped, or I-shaped. System (20) according to any of claims 1 to 5, characterized in that the cover (2) comprises at least one additional layer (22) on, between or under the first (8), second (10) and / or third (12) layers. System (20) according to any of the preceding claims, characterized in that the volume of the reaction chambers (6) of the bioassay cartridge (4) is between 5 µl and 500 µl, preferably between 5 µl and 50 µl. or between 50 µl and 500 µl, and most preferably between 10 µl and 30 µl.